1. Field of the Invention
The invention is drawn to molecular markers for mutant alleles of soybean associated with the low P34 allergen phenotypes.
2. Description of the Prior Art
Soybean is an important source of vegetable oil and high protein meal that is incorporated into many foods and feeds, and the use of soybean meal by the food industry is increasing. However, anti-nutritional compounds such as phytate, raffinose family oligosaccharides, and allergenic proteins that accumulate during normal soybean seed development limit the extent to which soybean meal can be included in livestock diet formulations. Livestock such as weanling pigs have been shown to have a sensitivity to soybean meal proteins (Li et al. 1991. Interrelationship between hypersensitivity to soybean proteins and growth performance in early-weaned pigs. J. Anim Sci. 69:4062-4069; and Li et al. 1990. Transient hypersensitivity to soybean meal in the early-weaned pig. J. Anim Sci. 68:1790-1799). For humans too, soybean seeds contain multiple proteins that are considered to be allergenic. The United States Food and Drug Administration recently recognized soybean as one of the major food allergens and new food allergen labeling requirements are in effect.
The dominant or major soybean allergen is Gly m Bd 30K, a papain superfamily cysteine protease-type protein which is also known as P34 (Kalinski et al. 1990. Molecular cloning of a protein associated with soybean seed oil bodies that is similar to thiol proteases of the papain family. J. Biol. Chem. 265:13843-13848; and Ogawa et al. 1993. Identification of the soybean allergenic protein, Gly m Bd 30K, with the soybean seed 34-kDa oil-body-associated protein. Biosci Biotechnol Biochem. 57:1030-3). This protein provoked a response in almost two thirds of patients examined in one study (Ogawa et al. 1991. Investigation of the IgE-binding proteins in soybeans by immunoblotting with the sera of the soybean-sensitive patients with a topic dermatitis. J Nutr Sci Vitaminol (Tokyo). 37:555-65). Although P34 is not an abundant seed protein, it is consistently present in nearly all germplasm accessions evaluated (Joseph et al. 2006. Evaluation of Glycine germplasm for nulls of the immunodominant allergen P34/Gly m Bd 30k. Crop Sci 46:1755-1763; Xu et al. 2007. Proteomic analysis of the distribution of the major seed allergens in wild, landrace, ancestral, and modern soybean genotypes. Journal of the Science of Food and Agriculture 87:2511-2518; and Yaklich et al. 1999. analysis of the distribution of the major soybean seed allergens in a core collection of Glycine max accessions. Crop Sci 39:1444-1447).
Transgenic suppression was successful in downregulation of P34 accumulation in soybean seeds with no apparent negative consequences to normal seed development and composition (Herman et al. 2003. Genetic modification removes an immunodominant allergen from soybean. Plant Physiol. 132:36-43). While the P34 protein failed to accumulate in transgenic seeds, there were no collateral alterations in other seed proteins as assessed by 2D gel electrophoresis. Additionally, the protein storage vacuoles (PSVs) in the P34 suppressed seeds were indistinguishable from those in the control seeds despite the fact that the P34 protein has been shown to accumulate in the PSVs (Herman et al., 2003, ibid).
Recently, large scale screening of approximately 16,000 soybean accessions from the USDA germplasm collections identified only two Glycine max lines with reduced P34 protein accumulation in seeds (Joseph et al., 2006, ibid, the contents of which are incorporated by reference herein). Glycine max low P34 soybean accessions, PI 567476 and PI 603570A, were characterized as having normal accumulation of seed proteins other than P34 (Joseph et al., 2006, ibid). While these two soybean accessions were shown to accumulate greatly reduced levels of the P34 protein in mature seeds, an understanding of the molecular genetic basis for this trait was lacking.